Biochemistry and molecular biology international最新文献

Anti-tumor active polysaccharide against Sarcoma 180 was isolated by DEAE-Sepharose CL-6B and Sepharose 4B column chromatography from the hot-water soluble fraction of the mycelium of liquid-cultured Agaricus blazei mill. This polysaccharide did not react with antibodies of anti-tumor polysaccharides such as lentinan, gliforan, and FIII-2-b which is one of anti-tumor polysaccharides from Agaricus blazei. Moreover, the analyses of 13C-NMR and GC-MS suggested that this polysaccharide was preliminarily glucomannan with a main chain of beta-1,2-linked D-mannopyranosyl residues and beta-D-glucopyranosyl-3-O-beta-D-glucopyranosyl residues as a side chain. This polysaccharide was completely different from the anti-tumor polysaccharide from fruiting body of Agaricus blazei, beta-1,6-glucan.

Because the expression of the isoforms of protein kinase C (PKC) in human basal keratinocytes is not understood, the expression of PKC isoforms were screened in specimens of epidermal tissue from postburn skin and the normal locations for skin grafts in patients with second or higher degrees of flame injury. The expression of individual isoform was determined by Western blot technique. Only PKC alpha and zeta were detected in the epidermal tissues of normal and postburn skin and translocation occurred in PKC alpha. Patients without antibiotic treatment after flame injury had higher expressions of PKC alpha and zeta. These findings indicate that the mechanisms of cellular differentiation and growth in postburn epidermal tissue may be related to the expression and translocation of PKC alpha induced by intra- and extracellular stimulation. These changes in PKC alpha further activate the DAG/PKC signal transduction pathways.

Plasma lipid profile and abdominal obesity have been associated with breast cancer risk, however published results have been inconsistent. To clarify these associations we studied lipid and lipoprotein alterations, obesity degree and body fat distribution, in 30 newly diagnosed breast cancer patients without treatment and 30 controls matched by age and menopausal status. Both pre and postmenopausal breast cancer patients presented higher body mass index, waist/hip ratio and insulin levels than their matched controls. An increase in triglycerides and a decrease in HDL-cholesterol, especially in the HDL2 subfraction, were observed in patients with breast cancer. Besides, HDL particle from these patients showed increased apo A1/HDL-cholesterol ratio. These alterations were correlated with waist/hip ratio. The association between lipoprotein alterations and abdominal obesity independent of menopausal status, in untreated newly diagnosed breast cancer patients is reported for the first time in this study.

The human CR1 is a single chain membrane glycoprotein that is a member of the group of regulators of the complement activation system. In order to clarify the regulatory mechanisms of human CR1 gene expression, the 5'-flanking region of the human CR1 gene was isolated and its promoter was characterized. The CR1 expression was found to be transcriptionally up-regulated in HL60 cells by stimulation with DMSO. The cloned CR1 gene promoter was sequenced and computer analyzed. The potential promoter region lacks a distinct TATA box sequence. The transcription initiation site was determined by primer extension and several possible regulatory elements for transcription were found in the promoter region.

The isolation and partial characterization of a serine protease with arginine ester hydrolase activity from Bothrops jararacussu snake venom are described. The purification procedure consisted of a gel filtration of the crude venom on Sephadex G-75 followed by an ion-exchange chromatography of the active fraction on DEAE-cellulose and a rechromatography on Bio-Rex 70 resin. The esterase fraction (DI-III), M(r) = 25,000 by SDS-PAGE, showed proteolytic activity on fibrinogen and casein. After 2 hr incubation, the A alpha and B beta chains of fibrinogen were intensely hydrolysed, while the gamma chain kept apparently intact, even after 20 hr of incubation. In spite of that, DI-III did not clot fibrinogen. DI-III induced edema in the rat paw. Although unable to release bradykinin, it induced contractions of the isolated rat uterus. DI-III did not catalyse the hydrolysis of bradykinin. Its arginine ester hydrolase activity was completely inhibited by diisopropyl fluorophosphate after 1 hr incubation, but not by phenylmethylsulfonyl fluoride under the same conditions.

The sodium hydrogen exchanger isoform, NHE-1 plays an important role in electrolyte and water homeostasis. These functions are compromised in pregnancies complicated with preeclampsia. At present it is not known whether NHE-1 expression is altered during preeclampsia. In the present study the placental level of NHE-1 protein was measured using immunoblotting. Since prostaglandins regulate the secretory and absorptive functions, the levels of prostaglandin E-2 as well as the expression of cyclooxygenase-1 and -2 were also estimated. The amount of NHE-1 protein and cyclooxygenase-2 was reduced in preeclamptic placentas, whereas the level of cyclooxygenase-1 remained unaltered. In contrast, prostaglandin E-2 concentration was higher in preeclampsia. Suppression of NHE-1 might render the placenta with impaired uptake of water and electrolytes and therefore may be involved in the pathogenesis of preeclampsia. While prostaglandin E-2 may play a role in preeclampsia, these findings discount the induction of cyclooxygenase-genes for this increase.

The interaction between acetylsalicylic acid (aspirin) and membrane was studied by NMR spectra. (1) NMR spectra showed acetylsalicylic acid did not insert into membrane; (2) 1H NMR spectrum recorded by spin-echo pulse sequence showed protons of the aromatic ring interacted with membrane; (3) the change of spin-lattice relaxation (T1) of 31P was ascribed to the association of acetylsalicylic acid to the polar head of lecithin; (4) the self-diffusion coefficient measured by pulsed field gradients NMR showed the mobility of acetylsalicylic acid was restricted by membrane and that acetylsalicylic acid changed membrane viscosity. Based on the results, the relationship between the interaction and the mechanism of the wide pharmacological functions of acetylsalicylic acid is discussed.

The polymerization of actin induced by dilution has previously been reported, where a 1000-fold molar excess of ATP over actin resulted when actin was diluted to 4.0 micrograms/ml in low salt buffer A (0.1 mM ATP, 0.1 mM CaCl2, 2 mM Tris-HCl, pH 8.0, 5 mM 2-mercaptoethanol, 1 mM NaN3). Filaments formed by the addition of ATP to a 1000-fold molar excess over actin in buffer B (0.1 mM CaCl2, 2 mM Tris-HCl, pH 8.0, 1 mM NaN3) were then separated by gel-filtration. When ATP was removed from these filaments using Dowex-1, depolymerization occurred. Thus, the reversible polymerization induced by the dilution of actin or by addition of ATP can be ascribed to the binding of ATP at the low affinity site of actin.

Osteopontin (OPN) is a secreted glycoprotein implicated in cell adhesion. It contains the arginine-glycine-asparatic acid (RGD) cell adhesive domain and the thrombin cleavage sequence. Although thrombin cleavage of OPN has been shown to be of physiological importance, the function of C-terminal OPN fragment cleaved by thrombin remains unknown. To determine its role, we performed cell adhesion assays using glutathione S-transferase-OPN fusion protein fragments and full-length OPN fusion protein. The N-terminal fragment containing RGD motif promoted enhanced adhesion of mouse and human fibroblasts by 2.9 and 2.8 folds in comparison with full-length OPN, respectively. The enhanced adhesion of both cells mediated by N-terminal fragment was significantly suppressed by addition of C-terminal fragment lacking RGD motif that has less cell adhesive property than full-length OPN. These results suggest that the C-terminal domain may play a pivotal role in regulating OPN functions by suppressing the RGD-dependent cell adhesion.

The effects of alpha-Hederin, a triterpenoid saponin which exists in some oriental herbs, on the expression of liver cytochrome P450s were examined in mice. The administration of alpha-Hederin to mice significantly decreased the hepatic content of P450 and the activities of microsomal ethoxyresorufin O-deethylase, methoxyresorufin O-demethylase, and aniline hydroxylase, representative activities of cytochrome-P4501A1, -P4501A2, and -P4502E1, respectively, in a dose- and time-dependent manner. However, pentoxyresorufin O-dealkylase, a representative activity of cytochrome P4502B1/2, was decreased to a lesser extent. alpha-Hederin also decreased inducible monooxygenase activities in the same manner. Suppressions of P450 isozyme expression occurred in alpha-Hederin treated hepatic microsomes, as determined by immunoblot analysis in a manner consistent with that of the enzyme activity levels. Levels of mRNA of P4501A1/2 and P4502B1/2 were also decreased by alpha-Hederin as shown by Northern blot analysis. In contrast, the level of P4502E1 mRNA in the liver of alpha-Hederin treated mice was unchanged. These results suggest that alpha-Hederin may act as a more specific suppressor for P4501A and P4502E1 than P4502B and that the suppression involves decreases in mRNA levels except in the case of P4502E1.